Transcriptomic differences in chromatin and cell cycle regulation in A549 cells after irradiation with carbon ions and X-rays.

INTRODUCTION: High-linear energy transfer (LET) radiation such as carbon ions exhibits greater biological effectiveness than conventional low-LET X-rays, but the transcriptional mechanisms underlying this advantage remain incompletely understood. We hypothesized that high-LET radiation induces a qualitatively different transcriptional response rather than simply amplifying low-LET signaling.

METHODS: A549 non-small cell lung cancer cells were exposed to equal physical doses (8 Gy) of X-rays or carbon ions (LET 73 keV/µm), and transcriptomic profiling was performed 4 h post-irradiation. Differential expression analysis was integrated with Hallmark pathway enrichment using gene set enrichment analysis (GSEA), over-representation analysis (ORA), and leading-edge gene interrogation to identify shared and LET-dependent gene expression regulation.

RESULTS: Both radiation modalities activated a conserved DNA damage response characterized by p53 signaling and apoptosis-related genes. In contrast, carbon ions selectively suppressed mitotic regulators including CENPE, KIF2C, PLK1, and BUB1, consistent with transcriptional disruption of the replication-segregation machinery. High-LET irradiation additionally enriched inflammatory and stress-associated pathways, including tumor necrosis factor (TNF), Nuclear Factor κB (NF-κB) and extracellular matrix and adhesion-related signatures annotated within the Hallmark epithelial-mesenchymal transition (EMT) gene set. Carbon ions also downregulated multiple core and linker histone genes, revealing a chromatin regulatory reprogramming signature although this may reflect modulation of mRNA stability linked to replication stress and cell-cycle progression. KRAS-associated gene networks were enriched under high-LET conditions, reflecting convergence of stress-responsive signaling.

DISCUSSION: At equal physical doses, high-LET carbon ion irradiation is associated with a transcriptional program distinct from that of low-LET X-rays, characterized by downregulation of mitotic and chromatin regulatory programs and selective engagement of stress-associated signaling networks. These findings provide mechanistic insight into LET-dependent radiobiology and suggest transcriptional pathway remodeling may contribute to the enhanced biological effectiveness of carbon ions.